(A) H3K27me3 peak profiles are comparable between young and aged animals. A scatter plot (top panel) of 331 peaks and a genome browser mini-view (bottom, left panel) illustrated that H3K27me3 had …
(A) Western blot (top) and quantification (bottom) show an increase of H3K27me3 with age in muscle tissues. (mean ±SD of three biological repeats; student t-test). Genotype: 5905. (B) Quantitative …
(A) PRCs-deficient animals have extended lifespan. H3K27me3 western blot (left), H3K27me3 quantification (middle), and lifespan curve (right) for PRC single mutants. PRC2 heterozygous mutants of …
(A) PRCs-deficient animals have extended lifespan. PRC2 heterozygous mutants of indicated genotype reduce H3K27me3 levels and extend lifespan (top of the dashed line); PRC1 heterozygous mutants of …
(A) Circos plot of the H3K27me3 epigenome illustrates peak profiles that are highly preserved with age and in PRC2 mutants. Black boxes and lines (innermost circle) represented common peak regions, …
(A) Relative quantification of different histone marks between WT and PRC2 mutants. Only H3K27me2/3 were selectively reduced in PRC2 mutants. Western blot was from head tissues of 3d-old male flies. …
(A) H3K27me3 modification increases with age in head. Scatter plot showed H3K27me3 levels for protein-coding genes (transcriptional start site to transcriptional termination sites as annotated in …
(A) Western blot (top) and quantification (bottom) show an increase of H3K27me3 with age in head tissues. (mean ±SD of three biological repeats; student t-test). Genotype: 5905. (B) The distribution …
(A) Venn diagram shows genes commonly changed in PRC2 single mutants with indicated genotype (left panel). Differential expression genes were computed using DESeq based on normalized count from …
(A) Number of genes upregulated in PRC2 mutants with indicated genotype. Differential expression genes were computed using DESeq based on normalized count from three biological replicates (cutoff: p<…
(A) Untargeted metabolics identifies that metabolites of the glycolytic pathway (red) are decreased with age. For each metabolite, the median from 10 biological repeats was used to calculate the …
(A) Heatmap of metabolites exhibiting differential levels of 3d, 30d WT and 30d PRC2 mutants. Data normalization using support vector regression algorithm. Metabolomics was from head tissues of 3d- …
(A) Schematic representation of the glucose flux experiment in adult aging flies. Briefly, flies at 3d (young) and 25d (aged) were switched from 12C-glucose to 13C-glucose food for a fixed 5days, …
(A) Glucose content shows a slight increase in PRC2 mutants, but this increase is not statistically significant (left panel). Pyruvate, one key end product of glycolysis, is increased (right panel). …
(A) Tpi deficiency significantly diminishes the longevity phenotype of PRC2 trans-heterozygous double mutants. (for lifespan assay: 25°C; n > 200 per genotype; log-rank test). Genotypes: Pclc421/+; S…
(A)-(D) and (F)-(I) Assessments of adult lifespan (A) and (F), oxidation test (B) and (G), climbing ability (C) and (H), and the levels of ATP and the ratio of NADH/NAD+ (D) and (I), show no …
(A) Tpi protein western blot (top) and quantification (bottom) shows a decrease with age. (mean ±SD of three biological repeats; student t-test). Western blot was from 3d- and 30d-old male flies. …
(A) and (B) Genomic structure and gene isoforms were shown according to the Flybase annotation. Color codes represented ATG (the translation start site), HA tag, CDS (coding sequence), 5’UTR, and …
Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
---|---|---|---|---|
Cell line (D. melanogaster) | S2R+ | This paper | FLYB:FBtc0000150 | |
Antibody | H3K27me3 (rabbit polyclonal) | Millipore | Cat#07–449; RRID:AB_310624 | |
Antibody | H3K27me2 (rabbit polyclonal) | Abcam | Cat#ab24684; RRID:AB_448222 | |
Antibody | H3K27me (rabbit polyclonal) | Millipore | Cat#07–448 | |
Antibody | H3K27ac (rabbit polyclonal) | Abcam | Cat#ab4729; RRID:AB_2118291 | |
Antibody | H3K9me3 (rabbit polyclonal) | Abcam | Cat#ab8898; RRID:AB_306848 | |
Antibody | H3K9ac (rabbit polyclonal) | Millipore | Cat#06–942; RRID:AB_310308 | |
Antibody | H3K23ac (rabbit polyclonal) | Millipore | Cat#07–355; RRID:AB_310546 | |
Antibody | H3K4me3 (rabbit polyclonal) | Millipore | Cat#07–473; RRID:AB_1977252 | |
Antibody | H3K4me2 (rabbit polyclonal) | Millipore | Cat#07–030; RRID:AB_310342 | |
Antibody | H3K4me (rabbit polyclonal) | Millipore | Cat#07–436; RRID:AB_310614 | |
Antibody | H3K36ac (rabbit polyclonal) | Millipore | Cat#07–540 | |
Antibody | H3K36me3 (rabbit polyclonal) | Abcam | Cat#ab9050; RRID:AB_306966 | |
Antibody | H3K14ac (rabbit polyclonal) | Millipore | Cat#07–353; RRID:AB_310545 | |
Antibody | H3K18ac (rabbit polyclonal) | Abcam | Cat#ab1191; RRID:AB_298692 | |
Antibody | H4K20me3 (rabbit polyclonal) | Abcam | Cat#ab9053 | |
Antibody | H3 (goat polyclonal) | Abcam | Cat#Ab12079 | |
Antibody | HA (rabbit monoclonal) | Cell signaling technology | Cat#3724 | |
Antibody | Tubulin (rabbit polyclonal) | Medical and biological laboratories co | Cat#PM054 | |
Antibody | Anti-Rabbit IgG | Sigma | Cat#A9169 | |
Antibody | Anti-Mouse IgG | Sigma | Cat#A4416 | |
Antibody | Anti-Goat IgG | Abcam | Cat#ab6471 | |
Recombinant DNA reagent | pBID UAS vector: HA-Tpi | Addgene | 35198 | |
Recombinant DNA reagent | pBID UAS vector: HA-Pgi | Addgene | 35198 | |
Recombinant DNA reagent | pBID vector: HA-Tpi genomic | Addgene | 35190 | |
Recombinant DNA reagent | pBID vector: HA-Pgi genomic | Addgene | 35190 | |
Recombinant DNA reagent | pU6b-sgRNA | Ren et al., 2013 | N/A | |
Commercial assay or kit | TURBO DNA-free kit | ThermoFisher | Cat#AM1907 | |
Commercial assay or kit | SuperSignal West Pico Chemilluminescent Subtrate | ThermoFisher | Cat#34078 | |
Commercial assay or kit | Nuclear-Cytosol Extraction Kit | Aogma | Cat#9988 | |
Commercial assay or kit | SYBR Select Master Mix | ThermoFisher | Cat#26161 | |
Commercial assay or kit | SuperScript III First-strand synthesis system for RT-PCR | ThermoFisher | Cat#18080–051 | |
Commercial assay or kit | ChIP-Grade Protein A/G Plus Agarose | ThermoFisher | Cat#26161 | |
Commercial assay or kit | QIAquick PCR Purification Kit | QIAGEN | Cat#28106 | |
Commercial assay or kit | Qubit dsDNA HS Assay Kit | ThermoFisher | Cat#Q32851 | |
Commercial assay or kit | High Sentivity DNA Analysis Kits | Agilent Technologies | Cat#5067–4626 | |
Commercial assay or kit | NEBNext DNA Library Prep Kit | New England Biolabs | Cat#E7370L | |
Commercial assay or kit | NEBNext Poly(A) mRNA Magnetic Isolation Module | New England Biolabs | Cat#E7490L | |
Commercial assay or kit | NEBNext Multiplex Oligos for Illumina | New England Biolabs | Cat#E7600S | |
Commercial assay or kit | NEBNext Ultra RNA library Prep Kit for Illumina | New England Biolabs | Cat#E7530L | |
Commercial assay or kit | RNase A/T1 Mix | ThermoFisher | Cat#EN0551 | |
Commercial assay or kit | Glucose (HK) Assay Kit | Sigma | Cat#GAHK20 | |
Commercial assay or kit | NAD/NADH Quantitation Kit | Sigma | Cat#MAK037 | |
Commercial assay or kit | NADP/NADPH Quantitation Kit | Sigma | Cat#MAK038 | |
Commercial assay or kit | ENLITEN ATP Assay System | Promega | Cat#FF2000 | |
Commercial assay or kit | Pyruvate Assay Kit | Sigma | Cat#: MAK071 | |
Commercial assay or kit | GSH/GSSG Ratio Detection Assay | Abcam | Cat#ab13881 | |
Commercial assay or kit | Amicon Ultra 0.5 mL centrifugal filters | Sigma | Cat#Z677108-96EA | |
Chemical compound, drug | Tris-buffer (1 mol/L, pH8.5) | Sangon Biotech | Cat#SD8141 | |
Chemical compound, drug | TRIZol Rreagent | ThermoFisher | Cat#15596018 | |
Chemical compound, drug | Chloroform | Sinopharm Chemical Reagent | Cat#10006818 | |
Chemical compound, drug | Isopropanol | Sinopharm Chemical Reagent | Cat#10006818 | |
Chemical compound, drug | DEPC-treated water | Invitrogen | Cat#46–2224 | |
Chemical compound, drug | NuPAGE 12% Bis-Tris Gel | ThermoFisher | Cat#NP0342BOX | |
Chemical compound, drug | PageRuler Prestained Protein Ladder | ThermoFisher | Cat#26616 | |
Chemical compound, drug | NuPAGE MOPS SDS Running Buffer | ThermoFisher | Cat#NP0001 | |
Chemical compound, drug | Immobilon Transfer Membranes | Millipore | Cat#: IPVH00010 | |
Chemical compound, drug | Glycine | beyotime | Cat#ST085-1000G | |
Chemical compound, drug | 20 X PBS | Sangon Biotech | Cat#B548117-0500 | |
Chemical compound, drug | 37% formaldehyde solution | Sigma | Cat#F1635 | |
Chemical compound, drug | Protease inhibitor cocktail tablets | Roche | Cat#11697498001 | |
Chemical compound, drug | RIPA buffer | Sigma | Cat#R-278–500 ML | |
Chemical compound, drug | 5M NaCl | ThermoFisher | Cat#AM9760G | |
Chemical compound, drug | 0.5M EDTA | ThermoFisher | Cat#15575–038 | |
Chemical compound, drug | 1M Tris-HCl | ThermoFisher | Cat#15568–025 | |
Chemical compound, drug | Triton X-100 | Sigma | Cat#T8787-50ML | |
Chemical compound, drug | 10% SDS | Sangon Biotech | Cat#SD8118 | |
Chemical compound, drug | RNase A/T1 Mix | ThermoFisher | Cat#EN0551 | |
Chemical compound, drug | Sodium dicarbonate | Sigma | Cat#S5761 | |
Chemical compound, drug | Proteinase K | Sangon Biotech | Cat#A100706 | |
Chemical compound, drug | 3M Sodium Acetate pH5.5 | ThermoFisher | Cat#AM9740 | |
Chemical compound, drug | Methyl viologen dichloride hydrate | Sigma | Cat#856177 | |
Chemical compound, drug | Hydrogen peroxide 30% | Sinopharm Chemical Reagent | Cat#10011208 | |
Chemical compound, drug | D(+)-Arabinose | Sangon Biotech | Cat#A600071-0100 | |
Chemical compound, drug | D-glucose | Sangon Biotech | Cat#A100188-0500 | |
Chemical compound, drug | Sodium hydroxide solution | Sigma | Cat#72068 | |
Chemical compound, drug | Methanol | Honeywell | Cat#LC230-2.5HC | |
Chemical compound, drug | Acetonitrile | Merck | Cat#1.00029.2500 | |
Chemical compound, drug | Water | Honeywell | Cat#LC365-2.5HC | |
Chemical compound, drug | Ammonium acetate | Sigma-Aldrich | Cat#73594–25 G-F | |
Chemical compound, drug | Ammonium hydroxide | Sigma-Aldrich | Cat#44273–100 mL-F | |
Chemical compound, drug | Sequencing Grade Modified Trypsin | Promega | Cat#V5111 | |
Chemical compound, drug | Centrifugal Filters Ultracel 30K | Amicon | Cat#UFC30SV00 | |
Chemical compound, drug | Water, LC/MS | J.T.Baker | Cat#9831–03 | |
Chemical compound, drug | Formic acid, eluent additive for LC-MS | Sigma | Cat#56302–50 ml-f | |
Chemical compound, drug | Acetonitrile,LC/MS,4L | J.T.Baker | Cat#9829–03 | |
Chemical compound, drug | Ammonium bicarbonate | Sigma | Cat#09830–1 KG | |
Chemical compound, drug | Urea | Sigma | Cat#U5128-500G | |
Chemical compound, drug | Trizma hydrochloride | Sigma | Cat#T5941-100G | |
Chemical compound, drug | Ammonium formate | Flucka | Cat#17843–50G | |
Chemical compound, drug | Lys(6) SILAC yeast | Silantes | Cat#234CXX-SYK6-509-01 | |
Chemical compound, drug | D-Glucose (U-13C6,99%) | Cambridge Isotope Laboratories, Inc. | Cat#110187-42-3 | |
Software, algorithm | Bowtie2 | Langmead and Salzberg (2012) | http://bowtie-bio.sourceforge.net/bowtie2/index.shtml | |
Software, algorithm | Samtools | Li et al. (2009) | http://samtools.sourceforge.net/; RRID:SCR_002105 | |
Software, algorithm | deepTools | Ramírez et al., 2014 | https://deeptools.github.io/ | |
Software, algorithm | homer | Heinz et al., 2010 | http://homer.ucsd.edu/homer/ngs/index.html; RRID:SCR_010881 | |
Software, algorithm | bwtool | Pohl and Beato, 2014 | https://users.dcc.uchile.cl/~peortega/bwtool/; RRID:SCR_003035 | |
Software, algorithm | Galaxy | Grüning et al., 2017 | https://github.com/bgruening/galaxytools; RRID:SCR_006281 | |
Software, algorithm | Cistrome | Liu et al., 2011 | http://cistrome.dfci.harvard.edu/ap/; RRID:SCR_000242 | |
Software, algorithm | BEDTools | Quinlan and Hall, 2010 | http://bedtools.readthedocs.io/; RRID:SCR_006646 | |
Software, algorithm | J-circos | An et al., 2015 | https://omictools.com/j-circos-tool; RRID:SCR_011798 | |
Software, algorithm | VENNY | Oliveros, 2007 | http://bioinfogp.cnb.csic.es/tools/venny/index.html | |
Software, algorithm | R | R Core Team, 2013 | https://www.r-project.org; RRID:SCR_001905 | |
Software, algorithm | DAVID Bioinformatics Resources | Huang et al., 2009b | https://david.ncifcrf.gov/; RRID:SCR_001881 | |
Software, algorithm | IGV | Robinson et al., 2011 | http://software.broadinstitute.org/software/igv/; RRID:SCR_011793 | |
Software, algorithm | STAR | Dobin et al., 2013 | https://github.com/alexdobin/STAR; RRID:SCR_005622 | |
Software, algorithm | HTSeq | Anders et al., 2015 | http://www-huber.embl.de/HTSeq/; RRID:SCR_005514 | |
Software, algorithm | DESeq | Anders and Huber, 2010 | http://www-huber.embl.de/users/anders/DESeq/; RRID:SCR_000154 | |
Software, algorithm | WGCNA | Langfelder and Horvath, 2008 | https://labs.genetics.ucla.edu/horvath/htdocs/CoexpressionNetwork/Rpackages/WGCNA/; RRID:SCR_003302 | |
Software, algorithm | XCMS | Smith et al., 2006 | http://www.bioconductor.org/packages/release/bioc/html/xcms.html; RRID:SCR_015538 | |
Software, algorithm | MetaboAnalyst 3.0 | Xia et al., 2015 | http://www.metaboanalyst.ca/; RRID:SCR_015539 | |
Software, algorithm | CAMERA | Kuhl et al. (2012) | https://bioconductor.org/packages/release/bioc/html/CAMERA.html; RRID:SCR_011924 | |
Software, algorithm | PD1.4 | Thermo | https://www.thermofisher.com/order/catalog/product/IQLAAEGABSFAKJMAUH | |
Software, algorithm | Pathways to PCDL (version B.07.00) | Agilent Technologies | ||
Software, algorithm | PCDL Manager (version B.07.00) | Agilent Technologies | ||
Software, algorithm | Profinder (version B.08.00) | Agilent Technologies | ||
Software, algorithm | Prism | GraphPad | v6; RRID:SCR_002798 |
Summary of H3K27me3 peak regions.
Summary of ChIP-seq experiments. Two criteria for peak calling: IP/input ≥ 2, signals spanning ≥3 kb. In total, 222 peak regions of H3K27me3 were reproducibly identified by four biological replicates at 3d. For each peak regions, information pertaining to chromosomal location and genes therein contained were given. ChIP-seq was using muscle tissues of 3d=old animals. Genotype: 5905.
Summary of CRISPR/Cas9-led gene mutagenesis and lifespan.
For each gene, two sgRNAs with indicated sequences, detailed deletion sites confirmed by Sanger sequencing, and PCR validation were shown. To name new CRISPR mutant, a superscript amended to the gene contained a letter c denoting CRISPR/Cas9-directed mutagenesis followed by the size of genomic deletion. Mutants were backcrossed (at least five generations) into 5905 (Flybase ID FBst0005905, w1118) to assure that phenotypes were not associated with any variation in background. Lifespan curves, together with 50% (median lifespan) and 10% survival data were listed. (for lifespan assay: 25°C; n ≥ 140 per genotype for curve; log-rank test).
Summary of transcriptomic analysis of PRC2 longevity mutants.
RNA-seq summary. This table summarizes five categories of genes and their transcriptional changes in long-lived PRC2 mutants of indicated genotype: (1) commonly upregulated and (2) downregulated genes in all PRC2 long-lived mutants; genes known to promote longevity upon upregulation, including (3) insulin pathway genes, (4) TOR signaling pathway genes, and (5) genes of miscellaneous pathways. RNA-seq was from muscle and head tissues of 30d-old male flies.
Summary of transcriptomic analysis of genes in glycolysis, citric acid cycle, pentose phosphorylation pathway, and oxidative phosphorylation.
This table summarizes metabolic genes and their transcriptional changes in long-lived PRC2 mutants of indicated genotype. RNA-seq was from head and muscle tissues of 3d- and 30d-old male flies.
Supplemental Materials and methods: Primer sequences used for the study.